TNF receptors have multiple roles in shaping adaptive and innate immune responses, and dysregulation of multiple TNF family members has been shown to be critical in the pathogenesis of many rheumatic diseases. The lab has two main interests: 1) Biology of TNF-family receptors in lymphocytes and mouse models of autoimmunity 2) Understanding the molecular pathogenesis of single-gene autoimmune and autoinflammatory diseases. Recently, the lab has focused on three members of the TNF-receptor superfamily.[unreadable] [unreadable] 1) Fas and Fas Ligand: The Fas receptor has been shown to be important in immunoreceptor-mediated apoptosis of activated T and B lymphocytes. Both humans and mice with germ line mutations in the death receptor Fas accumulate abnormal lymphocytes and develop systemic autoimmunity. While most patients with non-familial autoimmune disease do not carry germ line Fas mutations, there is evidence that Fas-mediated apoptosis may be impaired in the milieu of chronic inflammation. We have been investigating what signals regulate Fas-mediated apoptosis in T cells, with the eventual aim of harnessing these discoveries to modulate Fas-induced apoptosis for therapeutic goals in human disease. In activated CD4+ T cells, TCR restimulation triggers apoptosis that depends on interactions between the death receptor Fas and its ligand FasL. This process, termed restimulation-induced cell death (RICD), is a mechanism of peripheral immune tolerance. TCR signaling sensitizes activated T cells to Fas-mediated apoptosis, but what pathways mediate this process are not known. In the past year, we have identified the the RhoGTPases Rac1 and Rac2 as essential components in RICD. RNAi-mediated knockdown of Rac GTPases greatly reduced Fas-dependent TCR-induced apoptosis. The ability of Rac1 to sensitize T cells to Fas-induced apoptosis correlated with Rac-mediated cytoskeletal reorganization, dephosphorylation of ERM-family cytoskeletal linker proteins and translocation of Fas to lipid raft microdomains. In primary activated CD4+ T cells, Rac1 and Rac2 were independently required for maximal TCR-induced apoptosis. Activating Rac signaling may be a novel way to sensitize chronically stimulated lymphocytes to Fas-induced apoptosis, an important goal in the treatment of autoimmune diseases. [unreadable] In another project related to biology the biology of Fas and Fas Ligand, we are investigating whether defects in the RICD pathway may be part of the predisposition to autoimmune disease development in the Wiskott-Aldrich syndrome, a genetic immunodeficiency with an unusually high incidence of autoimmune complications. Wiskott-Aldrich syndrome (WAS) is a X-linked disorder caused by mutations in the WAS gene and affecting the expression of the WAS protein (WASp) Despite concurrent immunodeficiency, autoimmunity occurs in 40-70% of WAS patients. We have found that WASp-deficient mice older than 6 months produce autoantibodies, and develop proliferative glomerulonephritis with immune complex deposition. Activated CD4+ T lymphocytes from WASp-deficient mice had reduced apoptosis after restimulation through the TCR, due to a specific defect in Fas Ligand (FasL) secretion. These results suggest a novel role of WASp in FasL secretion and argue that WASp-deficient mice provide a good model for the study of autoimmune manifestations of WAS and the development of immunological and gene therapies for these complications. [unreadable] [unreadable] 2) TNF Receptor 1 (TNFR1). This receptor is critical for triggering inflammatory responses in myeloid cells of the immune system and other organ systems, and has been successfully targeted by biologic therapeutics in rheumatoid arthritis and other inflammatory diseases. We are working with the Genetics and Genomics Branch in NIAMS to understand the pathophysiology of inflammation in patients with the TNF Receptor Associated Periodic Syndrome (TRAPS) a genetic autoinflammatory disease associated with dominant mutations in TNFR1. We have recently discovered that TNFR1 mutant molecules associated with TRAPS are misfolded and accumulate in the endoplasmic reticulum. We are characterizing the mechanisms by which these mutant receptors lead to hyperactivity of the innate immune system. We have found that the intracellularly retained TNFR1 accumulates to abnormally high levels in TRAPS patients and knock-in mice harboring TRAPS-associated TNFR1 mutations. We are investigating the hypothesis that the intracellularly accumulated mutant receptors spontaneously signal in a TNF-independent manner, which promotes hyper-reactivity to inflammatory stimuli and the symptoms of TRAPS. It is hoped that blocking signaling pathways specifically hyperactivated in TRAPS will lead to therapeutic benefit in this difficult to treat periodic fever syndrome, and reveal new insights into the functions of TNFR1.[unreadable] [unreadable] 3) DR3: This TNF-family receptor highly related to TNFR1, also known as TRAMP, LARD, WSL-1, or TNFRSF25, is a death-domain containing tumor necrosis receptor primarily expressed in T cells. TL1A, the TNF-family ligand for DR3, can costimulate T cells, but the physiological function of TL1A-DR3 interactions in immune responses is not known. Using DR3-deficient mice, we have confirmed that DR3 is the critical receptor responsible for TL1A-induced T cell costimulation and dendritic cells as the likely source for TL1A during T cell priming. Despite its role in costimulation, DR3 is not required for T cell polarization into Th1, Th2 or Th17 effector subtypes or after priming with model antigens or Toxoplasma gondii. However, DR3 is required for immunopathology, local T cell accumulation and cytokine production in disease models that depend on diverse effector T cell subsets. DR3 is required for efficient T-cell infiltration and immunopathology in inflamed tissues and may be a promising therapeutic target for T-cell mediated autoimmune diseases.